Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies

Catharina H M J Van Elssen; Henrik Clausen; Wilfred T V Germeraad; Eric Paul Bennett; Paul P Menheere; Gerard M J Bos; Joris Vanderlocht

doi:10.1016/j.jim.2010.12.006

Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies

Catharina H M J Van Elssen, Henrik Clausen, Wilfred T V Germeraad, Eric Paul Bennett, Paul P Menheere, Gerard M J Bos, Joris Vanderlocht

11 Citations (Scopus)

Abstract

Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylation of MUC1, cancer-specific MUC1-Tn/STn antigens, which are highly immunogenic, become exposed. We aimed at developing a system that allows detection of antibodies directed to the native form of MUC1 and the underglycosylated MUC1-Tn epitopes. To this end, we made use of the Chinese Hamster Ovary (CHO) ldlD cell line stably transfected with MUC1. This cell line has a glycosylation defect, which can be reversed by addition of different monosaccharides to the cell culture and enables the production of cells expressing the MUC1-Tn glycoforms. After validation with glycospecific antibodies, the CHO-ldlD MUC1 system was used to detect serum MUC1 and MUC1-Tn antibodies. Using this system, we could confirm the presence of MUC1-Tn antibodies in the serum of a patient vaccinated with a truncated MUC1 peptide. This indicates that the CHO-ldlD MUC1 system represents a flow cytometry-based technique to detect antibodies binding to the underglycosylated MUC1 protein. This cellular system is complementary to the previously published methods to detect MUC1 serum antibodies, since the antibodies to the native protein are evaluated and therefore it can be effectively used for MUC1 antibody monitoring in vaccination studies as well as for functional assays.

Original language	English
Journal	Journal of Immunological Methods
Volume	365
Issue number	1-2
Pages (from-to)	87-94
Number of pages	8
ISSN	0022-1759
DOIs	https://doi.org/10.1016/j.jim.2010.12.006
Publication status	Published - 28 Feb 2011

Keywords

Amino Acid Sequence
Animals
Antibodies, Neoplasm
Antigens, Neoplasm
Breast Neoplasms
CHO Cells
Cancer Vaccines
Case-Control Studies
Cricetinae
Cricetulus
Female
Flow Cytometry
Glycosylation
Humans
Molecular Sequence Data
Mucin-1
Recombinant Proteins
Transfection
Vaccines, Subunit

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Access to Document

10.1016/j.jim.2010.12.006

Cite this

@article{beb55cd83901433dbe53f42ec8a3cd23,

title = "Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies",

abstract = "Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylation of MUC1, cancer-specific MUC1-Tn/STn antigens, which are highly immunogenic, become exposed. We aimed at developing a system that allows detection of antibodies directed to the native form of MUC1 and the underglycosylated MUC1-Tn epitopes. To this end, we made use of the Chinese Hamster Ovary (CHO) ldlD cell line stably transfected with MUC1. This cell line has a glycosylation defect, which can be reversed by addition of different monosaccharides to the cell culture and enables the production of cells expressing the MUC1-Tn glycoforms. After validation with glycospecific antibodies, the CHO-ldlD MUC1 system was used to detect serum MUC1 and MUC1-Tn antibodies. Using this system, we could confirm the presence of MUC1-Tn antibodies in the serum of a patient vaccinated with a truncated MUC1 peptide. This indicates that the CHO-ldlD MUC1 system represents a flow cytometry-based technique to detect antibodies binding to the underglycosylated MUC1 protein. This cellular system is complementary to the previously published methods to detect MUC1 serum antibodies, since the antibodies to the native protein are evaluated and therefore it can be effectively used for MUC1 antibody monitoring in vaccination studies as well as for functional assays.",

keywords = "Amino Acid Sequence, Animals, Antibodies, Neoplasm, Antigens, Neoplasm, Breast Neoplasms, CHO Cells, Cancer Vaccines, Case-Control Studies, Cricetinae, Cricetulus, Female, Flow Cytometry, Glycosylation, Humans, Molecular Sequence Data, Mucin-1, Recombinant Proteins, Transfection, Vaccines, Subunit",

author = "{Van Elssen}, {Catharina H M J} and Henrik Clausen and Germeraad, {Wilfred T V} and Bennett, {Eric Paul} and Menheere, {Paul P} and Bos, {Gerard M J} and Joris Vanderlocht",

year = "2011",

month = feb,

day = "28",

doi = "10.1016/j.jim.2010.12.006",

language = "English",

volume = "365",

pages = "87--94",

journal = "Journal of Immunological Methods",

issn = "0022-1759",

publisher = "Elsevier",

number = "1-2",

}

TY - JOUR

T1 - Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies

AU - Van Elssen, Catharina H M J

AU - Clausen, Henrik

AU - Germeraad, Wilfred T V

AU - Bennett, Eric Paul

AU - Menheere, Paul P

AU - Bos, Gerard M J

AU - Vanderlocht, Joris

PY - 2011/2/28

Y1 - 2011/2/28

N2 - Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylation of MUC1, cancer-specific MUC1-Tn/STn antigens, which are highly immunogenic, become exposed. We aimed at developing a system that allows detection of antibodies directed to the native form of MUC1 and the underglycosylated MUC1-Tn epitopes. To this end, we made use of the Chinese Hamster Ovary (CHO) ldlD cell line stably transfected with MUC1. This cell line has a glycosylation defect, which can be reversed by addition of different monosaccharides to the cell culture and enables the production of cells expressing the MUC1-Tn glycoforms. After validation with glycospecific antibodies, the CHO-ldlD MUC1 system was used to detect serum MUC1 and MUC1-Tn antibodies. Using this system, we could confirm the presence of MUC1-Tn antibodies in the serum of a patient vaccinated with a truncated MUC1 peptide. This indicates that the CHO-ldlD MUC1 system represents a flow cytometry-based technique to detect antibodies binding to the underglycosylated MUC1 protein. This cellular system is complementary to the previously published methods to detect MUC1 serum antibodies, since the antibodies to the native protein are evaluated and therefore it can be effectively used for MUC1 antibody monitoring in vaccination studies as well as for functional assays.

AB - Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylation of MUC1, cancer-specific MUC1-Tn/STn antigens, which are highly immunogenic, become exposed. We aimed at developing a system that allows detection of antibodies directed to the native form of MUC1 and the underglycosylated MUC1-Tn epitopes. To this end, we made use of the Chinese Hamster Ovary (CHO) ldlD cell line stably transfected with MUC1. This cell line has a glycosylation defect, which can be reversed by addition of different monosaccharides to the cell culture and enables the production of cells expressing the MUC1-Tn glycoforms. After validation with glycospecific antibodies, the CHO-ldlD MUC1 system was used to detect serum MUC1 and MUC1-Tn antibodies. Using this system, we could confirm the presence of MUC1-Tn antibodies in the serum of a patient vaccinated with a truncated MUC1 peptide. This indicates that the CHO-ldlD MUC1 system represents a flow cytometry-based technique to detect antibodies binding to the underglycosylated MUC1 protein. This cellular system is complementary to the previously published methods to detect MUC1 serum antibodies, since the antibodies to the native protein are evaluated and therefore it can be effectively used for MUC1 antibody monitoring in vaccination studies as well as for functional assays.

KW - Amino Acid Sequence

KW - Animals

KW - Antibodies, Neoplasm

KW - Antigens, Neoplasm

KW - Breast Neoplasms

KW - CHO Cells

KW - Cancer Vaccines

KW - Case-Control Studies

KW - Cricetinae

KW - Cricetulus

KW - Female

KW - Flow Cytometry

KW - Glycosylation

KW - Humans

KW - Molecular Sequence Data

KW - Mucin-1

KW - Recombinant Proteins

KW - Transfection

KW - Vaccines, Subunit

U2 - 10.1016/j.jim.2010.12.006

DO - 10.1016/j.jim.2010.12.006

M3 - Journal article

C2 - 21194532

SN - 0022-1759

VL - 365

SP - 87

EP - 94

JO - Journal of Immunological Methods

JF - Journal of Immunological Methods

IS - 1-2

ER -

Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies

Abstract

Keywords

UN SDGs

Access to Document

Fingerprint

Cite this